Our objective is to study nicotinic binding sites in rat brain to develop a model for study and to propose a functional role for nicotinic receptors in the central nervous system. It is believed by many that nicotine has an abuse liability and may be at least partially responsible for the chronic use of tobacco. In addition, smoking is known to alter the effectiveness of doses of certain substances such as diazepam. In addition, the natural isomer has been found to possess agonist/antagonist antinociceptive properties whereas the (plus)-isomer is inactive. This and other evidence suggest that nicotinic receptors exist in the central nervous system. In order to demonstrate the direct binding of nicotine to nicotinic receptors in brain tissue we synthesized previously unavailable radiolabeled (minus)-3H-nicotine with high specific activity (5 Ci/mmole). We also expect to prepare (plus)-3H-nicotine with a similar specific activity. We propose to locate these stereospecific binding sites by 1) comparing the regional distribution of (plus)- (minus)-3H-nicotine and 2) comparing their subcellular distribution. We also propose to analyze this binding in vitro, and then to study the displacement by known antagonists of nicotine such as mecamylamine, known metabolites such as cotinine and nornicotine and drugs such as diazepam. We also plan to initiate detailed agonist/antagonist studies and to investigate the effects of these isomers on synaptosomal release of norepinephrine and dopamine. Finally, we propose to provide ample supplies of previously unavailable (plus)-nicotine as a stable salt to other investigators in this area.